U. urealyticum is a common commensal of the urogenital tract, yet, it is an important cause of chorioamnion infection and morbidity and mortality in premature infants. Antigens important in the human immune response are poorly understood. There are 14 recognized serovars which can be divided into two biotypes. It has been postulated that only certain serovars or members of a single biotype may be able to cause invasive disease and that lack of serovar or biotype specific antibody may be an important risk factor. The present grant application is the first competitive renewal of a grant submitted in response to RFA 88-AI-12 the goal of which was to develop monoclonal antibodies (mAbs) to be used in epitope mapping to identify serovar or serogroup specific antigens. The investigator and colleagues previously identified an antigen complex on Uu, designated MB, which suggested that serovar- and/or serogroup specificity is at the epitope rather than the whole antigen level. During this grant period, they have shown the existence of MB on all 14 serovars; that this antigen contains both serovar and cross-reactive epitopes; that MB is produced not only in vitro but also in vivo; MB is one of the most predominant antigens recognized during Uu infection of humans; and mAbs to MB can protect against development of disease. By antibody-reactive peptide scanning (Pepscan), they have shown that the major immunoreactive site of MB serovar 3 as well as serovar specificity maps to the carboxy region. Using Pepscan analysis they have established a synthetic peptide enzyme immunoassay (EIA) which will allow them to determine the antibody reactivity to the most antigenic epitope(s) in sera of ureaplasma infected patients. Importantly, they have established that the unique structure of the carboxy domain allows mutations that result in size variation of the MB antigen. They have shown that the carboxy terminus is composed of concatemeric 6 amino acid repeats occurring in different clinical isolates in as few as 7 copies and as many as 42 copies. The investigator states that all indications are that this variation may be a critical determinant in production of disease. They have developed PCR primers which allow detection of size variants plus they have developed PCR techniques which are reliable as culture for detection of Uu in patient specimens. Specific aims of the present grant application are to: (1) clone and sequence MB from the remaining 13 Uu serovars using methods established in the present grant period; (2) develop PCR methods to detect serovars and MB size variants in vivo; and (3) establish by Pepscan the serovar and biotype specific epitopes of each of the remaining 13 serovars; and use a synthetic peptide-based EIA to determine the serovar, biogroup, and total MB antibody (including isotype and subclass) response in patients with documented invasive Uu infection. Antibody profiles will be compared to the serovar(s) and size variant(s) present in the affected site as determined by PCR. These studies will establish, characterize, and validate reagents which the investigators feel should prove useful in elucidating the role of different serovars and biotypes in production of invasive Uu disease and the role of serovar specific antibody in protection.